1. Field of the Invention
The genetic modification of higher plants by nontraditional genetic or plant breeding techniques has many possible applications. It may be desirable to modify plants to enhance existing plant properties such as yield, plant growth characteristics, tolerance to stress, disease resistance, etc. It may also be desirable to endow plants with new properties such as the ability to produce non-plant products of possible agricultural or pharmaceutical interest. Genetic and recombinant DNA technology has been successful in permitting the designed modification of certain bacteria (procaryotes) and lower organisms (eucaryotes), such as yeast. As yet, these techniques have not been successfully applied to higher plants for a number of reasons.
First, events which lead to desired genetic modifications in plants are usually infrequent. Microorganisms, unlike plants, can be rapidly multiplied under laboratory conditions allowing for the selection of infrequent genetic modification events. Such selection procedures in plants are, at best, long-term, bulky and expensive and, at worst, are completely undoable in the absence of an appropriate selection system. Second, routes have been found by which exogenous genetic information (DNA) can be readily introduced into many procaryotic microorganisms, but few such routes exist for plants. In many microorganisms, particularly in E. coli and other related bacteria, DNA can be readily introduced by transformation (introduction of DNA by itself) or by transduction (introduction of DNA via a bacteriophage or virus). In general, transformation and transduction procedures have not been described for higher plants. An exception is in the case of the tumor inducing (Ti) plasmid in the bacterium Agrobacterium. A portion of the Ti plasmid is transferred from bacterium to plant when Agrobacterium infects plants and produces a crown gall tumor. That type of DNA transfer has been recently utilized in certain genetic modification experiments involving plants. Third, much less is known about plant genes and how they operate than is known about the genes of bacteria and certain eucaryotic microorganisms.
The DNA of certain viruses such as cauliflower mosaic virus which infect plants may serve as "vehicles" for the introduction of "exogenous" or "foreign" DNA into plant cells. (The viral DNA is referred to as a "vehicle" and the exogenous or foreign DNA inserted into the vehicle is referred to as "passenger" DNA.) In using viral DNA as a vehicle, there are many considerations. First, if the viral DNA is to be propagated in the host plant in virion form (eventually encapsidated in virus particles), then any adaptation of the viral DNA necessary for the construction of a vehicle must not interfere with infectivity and movement of the virus around the plant, as occurs during the development of a systemic infection. Secondly, the passenger DNA must be inserted in the viral DNA vehicle so that the passenger DNA is stable as the virus multiplies and so that the passenger DNA is replicated and expressed (codes for a desired gene product) in the infected plant. Thirdly, any adaptation of the viral DNA should not prevent the efficient multiplication of the vehicle so that the "passenger" DNA is available in high copy number in infected plant cells.
It is therefore desirable to find ways in which plant viral DNA may be modified, while retaining infectivity, movement and the ability for high multiplication.
2. Description of the Prior Art
Howell et al, Science (1980) 208, 1265-1267 demonstrates the infectivity of cloned cauliflower mosaic virus correcting a previous report by Szeto et al, Science (1977) 196, 210-212 that cloned CaMV lost infectivity. Franck et al, (1980) Cell, 21, 285-294 describes the complete sequence of CaMV. Shalla et al, (1980) Virology 102, 381-388 describes the virus isolates CM1841 and CM4-184.